Uniform latex particles ("ULPs") are, in general, extremely uniform spheres of small diameter. Typical diameters range from less than about 0.1 .mu.m to about 100 .mu.m. Particles smaller than 5 .mu.m are usually prepared by emulsion polymerization. The result of this process is a series of particles with extremely uniform size distributions.
The principal use for ULPs is in the medical diagnostic area, wherein the particles are utilized for latex agglutination tests. Newer suggested uses include microbiological applications--e.g., bacterial typing and identification, and measurement of serum blood levels of antibiotics. (See, e.g., Bangs, L.B., "Uniform Latex Particles," presented at a workshop at the 41st National Meeting, American Association for Clinical Chemistry, 1989, and available in printed form from Seragen Diagnostics Inc., Indianapolis, IN; or Galloway, R.J., "Development of microparticle tests and immunoassays," .COPYRGT.1988 by Seradyn, Inc., Indianapolis, IN.) Other varieties of particles, such as amide-modified latex ("AML") and carboxylate-modified latex ("CML") have amide and carboxylic acid groups, respectively, on their surfaces. These functional groups permit covalent binding of antigens or antibodies to the surface of the ULPs for improved agglutination tests.
The largest use of ULPs is in the field of immunodiagnostics or immunoassays, especially in latex agglutination tests, where they are used to detect the presence of very minute amounts or concentrations or antigens or antibodies, in the blood, serum, urine, or cerebrospinal fluid. In essence, ULPs are used to magnify or visualize the antigen/antibody complex formation; they can also be used to quantify this reaction.
For example, if one is attempting to measure a particular antibody ("Ab"), an appropriate antigen ("Ag") is coated onto the latex particles. Since the Ab is divalent, it may bind to identical sites on two adjacent particles and link them together. Thus, if Ab present in an individual's sample is mixed with the Ag-coated particles, it will cause agglutination or coagulation of the particles; these aggregates are generally visible to the naked eye. This phenomenon is, essentially, the basis for latex agglutination tests ("LAT").
One major difficulty with LATs is the fact that the coated particles tend to spontaneously agglutinate. This is largely due to the fact that latex suspensions are colloidal suspensions of hydrophobic particles. The stability of the suspension is dependent upon the surface active charges; addition of small amounts of protein (approximately 10 .mu.g per mg of latex) can cause agglomeration, whereas continued addition of larger amounts of protein tends to increase particle stability.
This type of agglutination is also a problem in the chromatographic assays using colored or visible particles, such as the assay disclosed in published PCT Application No. W088/08534. In this assay, a sample is applied to a substrate of absorbent material, and analyte binds to antibody or antigen bearing, mobile colored latex particles. The particles to which analyte binds are themselves bound by immobilized immunoreagent as the sample chromatographically traverses the length of the absorbent material.
Various methods of addressing this problem and the related problem of nonspecific agglutination have been suggested, including the use of linkers and spacers. Some of the suggested spacers include Protein A, diaminoalkanes, naked antibodies, and streptavidin-biotin spacers, to name a few. The use of heterobifunctional linkers, halogen substituted carboxylic acids, bovine serum albumin, surfactants, and F(ab).sub.2 fragments has also been suggested. However, few of these suggestions prove entirely satisfactory, as they tend to interfere with the assay, many doing so in a manner that inhibits agglutination. This is, of course, completely unacceptable for LATs.
Therefore, in response to an express need for an immunoassay procedure with diverse applicability, which avoids the agglomeration problems of other assays and which promotes the goals of improved accuracy and greater resolution, and which is also elegant in its simplicity, the Applicants hereby disclose the present invention, including its equivalents thereof.